Cytotoxic T-lymphocytes (CTLs) are a critical component of effective human immune responses to tumors or viral infections. Cytotoxic T-lymphocytes destroy neoplastic cells or virus infected cells through recognition of antigenic peptides presented by MHC class I molecules on the surface of the affected target cells. These antigenic peptides are degradation products of foreign proteins present in the cytosol of the affected cell, which are processed and presented to CTLs through the endogenous MHC class I processing pathway.
Although the recognition of a foreign protein in the context of the MHC class I molecule may be sufficient for the recognition and destruction of affected target cells by CTLs, the induction of antigen-specific CTLs from T-lymphocyte precursors requires additional signals. Specialized antigen presenting cells (APCs) can provide both the antigen-MHC class I ligand and the accessory signals required in the induction phase of CTL-mediated immunity. General properties of APCs include MHC class I and class II expression, expression of various adhesion molecules important for APC-lymphocyte interaction, and expression of costimulatory molecules such as CD80 and CD86. Examples of APCs include macrophages and dendritic cells (including cutaneous epidermal Langerhans cells, dermal dendritic cells, and dendritic cells resident in lymph nodes and spleen).
Attempts to induce antigen-specific CTL responses in vivo by immunization with killed tumor cells, killed virus-infected cells, or component proteins have generally been unsuccessful, presumably because proteins in the extracellular fluids cannot enter the cytosol and access the MHC class I presentation pathway.
Mayordomo, et al., 1995, Nature Med. 1(12): 1297-1302 disclose in vitro culture of peptide-pulsed dendritic cells, which show protection against the associated tumor challenge. The authors state that dendritic cells cultured in the presence of GM-CSF+IL-4 and transfected with chicken ovalbumin (OVA) were capable of preventing establishment of an OVA.sup.+ tumor, but not the untransfected parental melanoma.
Porgador and Giboa, 1995, J. Exp. Med. 182: 255-260 discloses use of dendritic cells to induce CTLs. No tumor challenge data is presented and no mention of CTL-mediated cross-priming with a peptide antigen is presented.
Nabel and coworkers (1995, Annals of the NY Academy of Sciences 772: 227-31; Human Gene Therapy, 1994, 5(1):57-77; Proc. Natl. Acad. Sci., 1993, 90:11307-11311) disclose a method of cancer immunotherapy whereby a known tumor antigen is delivered to tumor cells in vivo to stimulate cell-mediated immunity against tumor growth. The authors do not disclose immunization with a generic artificial target antigen to achieve cross-priming either in vitro or in vivo.
Boon, 1992, Advances in Cancer Research 58:177-210, reviews the field of tumor rejection antigens as it stood in 1992. The author offers a perspective in regard to the use of tumor rejection antigens regarding methods of tumor immunization. However, the author does not suggest or teach immunization with a generic artificial target antigen to achieve cross-priming either in vitro or in vivo.
Wahl, et al., 1995, J. Immunotherapy with Emphasis on Tumor Immunology 17(1): 1-11, show increased enhanced generation of therapeutic T-cells in response to transfection of MHC class I gene H-2Ks to a poorly immunogenic tumor cell line. The authors do not address immunization by a generic artificial target antigen to achieve cross-priming either in vitro or in vivo.
Sato, et al., 1995, Clinical Immunology & Immunopathology, 74(1):35-43, studied the immune response to dinitrophenyl-modified tumor cells. The authors detected an inflammatory response to patient immunization with these modified cells along with tumor regression. Again, the authors do not address immunization by a generic artificial target antigen.
Flamand, et al., 1994, Eur. J. Immunol. 24: 605-610 disclose in vitro culture of dendritic cells, pulsed with a peptide antigen, and subsequent induction of a T-cell dependent humoral response to the B cell tumor BCL1. No mention of cross-priming with ATA targeted CTLs is taught or suggested.
Williams, et al. (1991, Proc. Natl. Acad. Sci. USA 88: 2726-2730) showed the expression of the protein luciferase in intact epidermal cells following biolistic (biobalistic) delivery of the firefly luciferase gene.
Tang, et al. (1992, Nature 356: 152-154) utilized a biolistic (biobalistic) device to produce a humoral response to a foreign protein. A gene encoding hGH under control of either the CMV promoter or the .beta.-actin promoter was delivered to the epidermal tissue of mice. Anti-hGH antibodies were detected in mice in response to this immunization procedure.
Fynan, et al. (1993, Proc. Natl. Acad. Sci. USA 90: 11478-11482) confirmed the findings of Tang, et al. by using a plasmid DNA construct encoding an influenza virus hemagglutinin glycoprotein. Fynan, et al. compared humoral responses generated by gene gun delivery of DNA coated gold beads to the epidermis with other mechanisms and found that the use of a biolistic (biobalistic) device 1) resulted in 95% protection to a lethal influenza challenge, 2) was the most efficient route for DNA immunization, proving to be substantially more effective than mucosal, intramuscular, or intravenous administration, and, 3) required 250 to 2500 times less DNA than saline inoculations. Direct targeting of APC cells for genetic immunization is not disclosed or suggested by Fynan, et al.
Liu and colleagues (Montgomery et al., 1993, DNA Cell Biol. 12:777-783; Ulmer et al., 1993, Science. 259:1745-1749; Donnelly et al., 1995, Nature Medicine 1:583-587) have demonstrated that untargeted, nonspecific intramuscular injection of naked DNA induces antigen-specific CTL responses to viral proteins and protective immunity to viral challenge.
Sun, et al. (1995, Proc. Natl. Acad. Sci. USA 92: 2889-2893) utilized a biolistic (biobalistic) device to produce an anti-tumor response in mice. The authors delivered a plasmid construct expressing IL-6 directly to a tumor site in mice. Expression of IL-6 afforded a form of cytokine gene therapy nonspecifically directed at the tumor.
Kundig et al. (1995, Science. 268:1343-1346) demonstrate that protein antigen localization to the lymphoid organs is critical for the induction of antigen-specific CTL responses in vivo.
Kovacsovics-Bankowski and Rock (1995, Science 267: 243-246) demonstrate a phagosome-to-cytosol pathway for protein antigens not normally presented through the MHC class I endogenous pathway. The authors speculate that proteins in particulate form internalized within phagosomes are in fact able to enter the cytosolic pathway for MHC class I presentation.
Despite the efforts documented in the above reference material, there remains a need to develop a cancer immunotherapy procedure which stimulates protective and therapeutic immunity to a wide variety of tumor types. The present invention both addresses and meets this need.